Biology subjects

Shy, M. E.

Publications and source records attributed to Shy, M. E..

4 recordsLinked to original sources

bioRxiv2024

Activation of XBP1s attenuates disease severity in models of proteotoxic Charcot-Marie-Tooth type 1B

Mutations in myelin protein zero (MPZ) are generally associated with Charcot-Marie-Tooth type 1B (CMT1B) disease, one of the most common forms of demyelinating neuropathy. Pathogenesis of some MPZ mutants, such as S63del and R98C, involves the misfolding and retention of MPZ in the endoplasmic reticulum (ER) of myelinating Schwann cells. To cope with proteotoxic ER-stress, Schwann cells mount an unfolded protein response (UPR) characterized by activation of the PERK, ATF6 and IRE1/XBP1 pathways. Previous results showed that targeting the PERK UPR pathway mitigates neuropathy in mouse models of CMT1B; however, the contributions of other UPR pathways in disease pathogenesis remains poorly understood. Here, we probe the importance of the IRE1/XBP1 signalling during normal myelination and in CMT1B. In response to ER stress, IRE1 is activated to stimulate the non-canonical splicing of Xbp1 mRNA to generate spliced Xbp1 (Xbp1s). This results in the increased expression of the adaptive transcription factor XBP1s, which regulates the expression of genes involved in diverse pathways including ER proteostasis. We generated mouse models where Xbp1 is deleted specifically in Schwann cells, preventing XBP1s activation in these cells. We observed that Xbp1 is dispensable for normal developmental myelination, myelin maintenance and remyelination after injury. However, Xbp1 deletion dramatically worsens the hypomyelination and the electrophysiological and locomotor parameters observed in young and adult CMT1B neuropathic animals. RNAseq analysis suggested that XBP1s exerts its adaptive function in CMT1B mouse models in large part via the induction of ER proteostasis genes. Accordingly, the exacerbation of the neuropathy in Xbp1 deficient mice was accompanied by upregulation of ER-stress pathways and of IRE1-mediated RIDD signaling in Schwann cells, suggesting that the activation of XBP1s via IRE1 plays a critical role in limiting mutant protein toxicity and that this toxicity cannot be compensated by other stress responses. Schwann cell specific overexpression of XBP1s partially re-established Schwann cell proteostasis and attenuated CMT1B severity in both the S63del and R98C mouse models. In addition, the selective, pharmacologic activation of IRE1/XBP1 signaling ameliorated myelination in S63del dorsal root ganglia explants. Collectively, these data show that XBP1 has an essential adaptive role in different models of proteotoxic CMT1B neuropathy and suggest that activation of the IRE1/XBP1 pathway may represent a therapeutic avenue in CMT1B and possibly for other neuropathies characterized by UPR activation.

neuroscience↗

bioRxiv2023

Structural context of homomeric interactions in the Ig domain of the MPZ (P0) myelin adhesion protein and relation to Charcot-Marie-Tooth disease phenotype variants

Mutations in Myelin Protein Zero (MPZ) account for 5% of Charcot-Marie-Tooth cases and can cause demyelinating or axonal phenotypes, reflecting the diverse roles of MPZ in Schwann cells. MPZ holds the apposing membranes of the myelin sheath together, with the adhesion role fulfilled by the extracellular lmmunoglobulin-like domain (lgMPZ), which can oligomerize. Current knowledge for how the lgMPZ might form oligomeric assemblies involving 3 weakly-interacting interfaces has been extrapolated from a protein crystal structure in which individual rat lgMPZ subunits are packed together under artificial conditions. These interfaces include one that organizes the lgMPZ into tetramers, a dimer interface that could link tetramers together, and a third hydrophobic interface that could mediate binding to lipid bilayers or the same hydrophobic surface on another lgMPZ domain. There are at present no data confirming whether the proposed lgMPZ interfaces actually mediate oligomerization in solution, whether they are required for the adhesion activity of MPZ, whether they are important for myelination, or whether their loss results in disease. We performed NMR and SAXS analysis of wild-type lgMPZ as well as mutant forms with amino-acid substitutions designed to interrupt its presumptive oligomerization interfaces. Here, we confirm the interface that mediates lgMPZ tetramerization, but find that dimerization is mediated by a distinct interface that has yet to be identified. We next correlated CMT phenotypes to subregions within lgMPZ tetramers. Axonal late-onset disease phenotypes (CMT2l/J) map to surface residues of lgMPZ proximal to the transmembrane domain. Early-onset demyelinating disease phenotypes (CMT1B/Dejerine-Sottas syndrome) map to two groups: one is described by variants that disrupt the stability of the lg-fold itself and are largely located within the core of the lg domain; whereas another describes a surface on the distal outer surface of lgMPZ tetramers. Computational docking studies predict that this latter disease-relevant subregion may mediate dimerization of lgMPZ tetramers.

neuroscience↗

bioRxiv2022

A humanized yeast model reveals dominant-negative properties of neuropathy-associated alanyl-tRNA synthetase mutations

Aminoacyl-tRNA synthetases (ARSs) are ubiquitously expressed, essential enzymes that ligate tRNA molecules to their cognate amino acids. Heterozygosity for missense variants or small in-frame deletions in five ARS genes causes axonal peripheral neuropathy, a disorder characterized by impaired neuronal function in the distal extremities. These variants reduce enzyme activity without significantly decreasing protein levels and reside in genes encoding homo-dimeric enzymes. These observations raise the possibility of a dominant-negative effect, in which non-functional mutant ARS subunits dimerize with wild-type ARS subunits and reduce overall ARS activity below 50%, breaching a threshold required for peripheral nerve axons. To test for these dominant-negative properties, we developed a humanized yeast assay to co-express pathogenic human alanyl-tRNA synthetase (AARS1) mutations with wild-type human AARS1. We show that multiple loss-of-function, pathogenic AARS1 variants repress yeast growth in the presence of wild-type human AARS1. This growth defect is rescued when these variants are placed in cis with a mutation that reduces dimerization with the wild-type subunit, demonstrating that the interaction between mutant AARS1 and wild-type AARS1 is responsible for the repressed growth. This demonstrates that neuropathy-associated AARS1 variants exert a dominant-negative effect, which supports a common, loss-of-function mechanism for ARS-mediated dominant peripheral neuropathy.

genetics↗

bioRxiv2021

Treatment with IFB-088 improves neuropathy in CMT1A and CMT1B mice

Charcot Marie Tooth diseases type 1A (CMT1A), caused by duplication of Peripheral Myelin Protein 22 (PMP22) gene, and CMT1B, caused by mutations in myelin protein zero (MPZ) gene are the two most common forms of demyelinating CMT (CMT1) and no treatments are available for either. Prior studies of the MpzSer63del mouse model of CMT1B have demonstrated that protein misfolding, endoplasmic reticulum (ER) retention and activation of the unfolded protein response (UPR) contributed to the neuropathy. Heterozygous patients with an arginine to cysteine mutation in MPZ (MPZR98C) develop a severe infantile form of CMT1B which is modeled by MpzR98C/+ mice that also show ER-stress and an activated UPR. C3-PMP22 mice are considered to effectively model CMT1A. Altered proteostasis, ER-stress and activation of the UPR have been demonstrated in mice carrying Pmp22 mutations. To determine whether enabling the ER-stress/UPR and readjusting protein homeostasis would effectively treat these models of CMT1B and CMT1A we administered Sephin1/IFB-088/icerguestat, a UPR modulator which showed efficacy in the MpzS63del model of CMT1B, to heterozygous MpzR98C and C3-PMP22 mice. Mice were analyzed by behavioral, neurophysiological, morphological and biochemical measures. Both MpzR98C/+ and C3-PMP22 mice improved in motor function and neurophysiology. Myelination, as demonstrated by g-ratios and myelin thickness, improved in CMT1B and CMT1A mice and markers of UPR activation returned towards wild type values. Taken together our results demonstrate the capability of IFB-088 to treat a second mouse model of CMT1B and a mouse model of CMT1A, the most common form of CMT. Given the recent benefits of IFB-088 treatment in Amyotrophic Lateral Sclerosis and Multiple Sclerosis animal models, these data demonstrate its potential in managing UPR and ER-stress for multiple mutations in CMT1 as well as in other neurodegenerative diseases.

neuroscience↗